Effects of Ca2+-ATPase inhibitors, ionomycin, and pharmacological modulators of ryanodine receptor on calcium release from intracellular pools and onoscillatory contractile behavior in Physarum polycephalum

Changes in calcium levels in organelles of the plasmodium of the myxomycete Physarum polycephalum were analyzed using the fluorescent calcium indicato r chlortetracycline (CTC). Both the Ca2+-ATPase inhibitor 2,5'-di(terlbutyl )-1,4-benzohydroquinone (BHQ) (100 mu M) and the calcium ionophore ionomyci n (1 mu M) induce a significant decrease in fluorescence level (by 30%) in CTC-stained microplasmodia; this is caused by release of calcium from intra cellular storage compartments. An activator of ryanodine receptors, caffein e (10-50 mM), is less effective on Ca2+ release than BHQ or ionomycin, and their inhibitor, ryanodine (100 mu M), almost completely blocks the respons e to caffeine, but only slightly decreases the effects of BHQ or ionomycin. Procaine, another inhibitor of ryanodine receptors, at 10 mM concentration completely abolishes both the BHQ and the ionomycin responses, but 50 mM i s necessary to block the effect of 25 mM caffeine. These results suggest th at both the BHQ- and the ionomycin-dependent Ca2+ releases occur through th e ryanodine receptor and are to be considered as calcium-induced Ca2+ relea se (CICR). Both the ionomycin and the BHQ responses persist in the presence of Cd2+, which blocks Ca2+ channels of the plasmalemma. In most cases, Cd2 + itself induces release of Ca2+ from the CTC-stained calcium pool; the mor e effective Cd2+ is, the less the following ionomycin or BHQ responses occu r. This indicates that Ca2+ entry through plasmalemma plays no significant role in the ionomycin- or BHQ-evoked initiation of CICR, and that the Cd2+- and BHQ/ionomycin-depleted Ca2+ stores overlap.